CelLytic™ NuCLEAR™ Extraction Kit

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	- Nuclear and cytoplasmic extracts are prepared with a SIGMA CelLytic NuCLEAR Extraction Kit, utilizing hypotonic lysis buffer and IGEPAL (NXTRACT, SIGMA).

Protocol tips
- Nuclear and cytoplasmic extracts are prepared with a SIGMA CelLytic NuCLEAR Extraction Kit, utilizing hypotonic lysis buffer and IGEPAL (NXTRACT, SIGMA).

Publication protocol

ARPE-19 cells grown to confluency in 10 cm plates were pretreated with media (DMEM/F-12/10% FBS), 20 μM TLR9 control oligonucleotide in media, or 20 μM TLR9 inhibitory oligonucleotide in media for 2 hours at 37°C. Plates, except uninfected control plate, were infected with HSV-1 KOS at a MOI of 5 for 6 hours at 37°C. Cells were washed twice with cold PBS, collected by scraping, and pelleted at 1000 rpm for 10 minutes at 4°C in a Beckman Model TJ-6 table top centrifuge. Nuclear and cytoplasmic extracts were prepared with a SIGMA CelLytic NuCLEAR Extraction Kit, utilizing hypotonic lysis buffer and IGEPAL (NXTRACT, SIGMA). The protein concentration of each extract was determined by BCA assay (Pierce, Rockford, IL, 23225).

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Manufacturer protocol

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